LOCUS       sequence-466788-        7327 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  2,4-Diacetylphloroglucinol (DAPG)-regulatable EGFP expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7327)
  AUTHORS   Tominaga M, Shima Y, Nozaki K, Ito Y, Someda M, Shoya Y, Hashii N, 
            Obata C, Matsumoto-Kitano M, Suematsu K, Matsukawa T, Hosoya K, 
            Hashiba N, Kondo A, Ishii J
  TITLE     Designing strong inducible synthetic promoters in yeasts.
  JOURNAL   Nat Commun. 2024 Dec 19;15(1):10653. doi: 
            10.1038/s41467-024-54865-z.
  PUBMED    39702268
REFERENCE   2  (bases 1 to 7327)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Commun."; date: "2024-12-19"; volume: "15(1):10653. doi"; pages: " 
            10.1038/s41467-024-54865-z"
COMMENT     Sequence Label: pMT949
FEATURES             Location/Qualifiers
     source          1..7327
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(10..28)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        96..200
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(201..269,270..1061)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(419..438)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1232..1820
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1721..1740
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1974..1991
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    2108..2129
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(2144..2149,2150..2167,2168..2174)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2182..2198
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     2187..2209
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     2206..2222
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2206..2222
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(4157..4159,4160..4162,4163..4873)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     terminator      4951..5197
                     /gene="Pichia pastoris AOX1"
                     /label=AOX1 terminator
                     /note="transcription terminator for AOX1"
     CDS             complement(5846..6640)
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(5957..5976)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     6567..6586
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     complement(6933..6950)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(6933..6949)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6942..6964)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(7158..7177)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7277..7299
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
ORIGIN
        1 gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt
       61 cttagacgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt
      121 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat
      181 aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt
      241 ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg
      301 ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga
      361 tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc
      421 tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac
      481 actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg
      541 gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca
      601 acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg
      661 gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg
      721 acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg
      781 gcgaactact tactctagct tcccggcaac aattaataga ctggatggag gcggataaag
      841 ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg
      901 gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct
      961 cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac
     1021 agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac caagtttact
     1081 catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga
     1141 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
     1201 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     1261 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     1321 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc
     1381 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     1441 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     1501 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     1561 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     1621 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     1681 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
     1741 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
     1801 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
     1861 gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta
     1921 ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag cgcagcgagt
     1981 cagtgagcga ggaagcggaa gagcgcccaa tacgcaaacc gcctctcccc gcgcgttggc
     2041 cgattcatta atgcagctgg cacgacaggt ttcccgactg gaaagcgggc agtgagcgca
     2101 acgcaattaa tgtgagttag ctcactcatt aggcacccca ggctttacac tttatgcttc
     2161 cggctcgtat gttgtgtgga attgtgagcg gataacaatt tcacacagga aacagctatg
     2221 accatgatta cgccaagctt gcggccgcgg atccatggtt gtgcgacaga ttcactgtga
     2281 aagactgttc attataccca cgtttcactg ggagatgtaa gccttaggtg ttttaccctg
     2341 attagataat acaataacca acagaaatac gagaatctaa actaatttcg atgattcatt
     2401 tttcttttta ccgcgctgcc tcttttggca attctttcac ctatattcta ccttctcttt
     2461 ccttttgttc taaacttatt accagctatc tatgtcgaat caagaagaag gacttaaact
     2521 gtggggtggc aggtttactg gggctactga ccccttgatg gatttgtata acgcttcctt
     2581 accttacgac aagaaaatgt acaaggtgga tttagaagga acaaaagttt acactgaggg
     2641 cctggagaaa attaatttgc taactaaaga cgaactaagt gagattcatc gtggtctcaa
     2701 attgattgaa gcagagtggg cagaagggaa gtttgttgag aagccagggg atgaggatat
     2761 tcacactgct aatgaacgtc gcttgggtga gttgattggt cgtggaatct ctggtaaggt
     2821 tcataccgga aggtctagaa atgatcaagt tgccactgat atgcggttgt atgtcagaga
     2881 caatctaact cagttggctg actatctgaa gcagttcatt caagtaatca tcaagagagc
     2941 tgaacaggaa atagacgtct tgatgcccgg ttatactcac ttgcaaagag ctcaaccaat
     3001 cagatggtct cactggttga gcatgtatgc tacctatttc actgaagatt atgagagact
     3061 gaatcaaatc gttaaaaggt tgaacaaatc cccattggga gctggagctt tggctggtca
     3121 tccttatgga attgatcgtg aatacattgc tgagagatta gggtttgatt ctgttattgg
     3181 taattctttg gccgctgttt cagacagaga ttttgtagtc gaaaccatgt tctggtcttc
     3241 gttgtttatg aatcatattt ctcgattctc agaagatttg atcatttact ccactggaga
     3301 gtttggattt atcaagttgg cagatgctta ttctactgga tcttctctga tgcctcaaaa
     3361 aaaaaaccca gactctttgg agttattgag gggtaaatct ggtagatgtt ttggggcctt
     3421 ggctggtttc ctcatgtcta ttaagtccat tccgtcaacc tataacaaag atatgcaaga
     3481 ggataaggag cctttatttg atactctaat cactgtagag cactcgattt tgatagcatc
     3541 cggtgtagtt tctaccttga acattgatgc cgaacgaatg aagaatgctc taactatgga
     3601 tatgctggct acagatcttg ccgactattt agttagaagg ggagttccat tcagagaaac
     3661 tcaccacatt tctggtgaat gtgtcagaca agccgaggag ttgaaccttt ctggtattga
     3721 tcagttgtcc ctcgaacaat tgaaatccat tgactcccgt tttgaggctg atgtggcttc
     3781 aacgtttgac tttgaagcca gtgttgaaaa aagaactgcc accggaggaa cttctaagac
     3841 tgctgtttta aagcaattgg atgcactgaa tgaaaagcta gagtcttgac tgcaggtcga
     3901 ctctagagga tcctatgtat gatacgaaac gtaccgtatc gttaaggtag cgtcaaaatt
     3961 taactgttct aacccctact tgacagcaat atataaacag aaggaagctg ccctgtctta
     4021 aacctttttt tttatcatca ttattagctt actttcataa ttgcgactgg ttccaattga
     4081 caagcttttg attttaacga cttttaacga caacttgaga agatcaaaaa acaactaatt
     4141 attcgaaacg actagtatgg tttctaaggg tgaagagttg ttcactggtg ttgttccaat
     4201 cttggttgag ttggacggtg acgttaacgg acacaagttc tctgtttctg gtgaaggtga
     4261 gggtgacgct acttacggaa agttgacttt gaagttcatc tgtactactg gtaagttgcc
     4321 agttccatgg ccaactttgg ttactacttt gacttacggt gttcagtgtt tctccagata
     4381 cccagaccac atgaagcagc acgatttctt caagtctgct atgccagagg gttacgttca
     4441 agagagaact atcttcttca aggacgacgg taactacaag actagagctg aggttaagtt
     4501 cgagggtgac acattggtta acagaatcga gttgaagggt atcgacttca aagaggacgg
     4561 aaacatcttg ggtcacaagt tggagtacaa ctacaactcc cacaacgttt acatcatggc
     4621 tgacaagcag aagaacggta tcaaggttaa cttcaagatc agacacaaca tcgaggacgg
     4681 ttccgttcaa ttggctgacc actaccaaca gaacactcca attggtgacg gtccagtttt
     4741 gttgccagac aaccactact tgtccactca atccgctttg tccaaggacc caaacgagaa
     4801 gagagatcac atggttttgt tggagttcgt tactgctgct ggtatcactt tgggtatgga
     4861 cgagttgtac aagtaactcg agccttagac atgactgttc ctcagttcaa gttgggcact
     4921 tacgagaaga ccggtcttgc tagattctaa tcaagaggat gtcagaatgc catttgcctg
     4981 agagatgcag gcttcatttt tgatactttt ttatttgtaa cctatatagt ataggatttt
     5041 ttttgtcatt ttgtttcttc tcgtacgagc ttgctcctga tcagcctatc tcgcagctga
     5101 tgaatatctt gtggtagggg tttgggaaaa tcattcgagt ttgatgtttt tcttggtatt
     5161 tcccactcct cttcagagta cagaagatta agtgagacgt tcgtttgtgc aagcttagat
     5221 cttctagaga caataagaag aaaaaaaaag aaaagcggtg ggggagggat tattaaataa
     5281 ggattatgta accccagggt accgttctat acatatttaa ggattattta ggacaatcga
     5341 tgaaatcggc atcaaactgg atgggagtat agtgtccgga taatcggata aatcatcttg
     5401 cgaggagccg cttggttggt tggtgagagg agtgaaatat gtgtctcctc acccaagaat
     5461 cgcgatatca gcaccctgtg ggggacacta ttggcctccc tcccaaacct tcgatgtggt
     5521 agtgctttat tatattgatt acattgatta catagctaaa ccctgcctgg ttgcaagttg
     5581 agctccgaat tccaatatta gtaaaatgcc tgcaagataa cctcggtatg gcgtccgacc
     5641 ccgcttaatt attttaactc ctttccaacg aggacttcgt aatttttgat tagggagttg
     5701 agaaacgggg ggtcttgata cctcctcgat ttcagatccc accccctctc agtcccaagt
     5761 gggacccccc tcggccgtga aatgcgcgca ctttagtttt tttcgcatgt aaacgccggt
     5821 gtccgtcatc tagagaattc cccgctcaga agaactcgtc aagaaggcga tagaaggcga
     5881 tgcgctgcga atcgggagcg gcgataccgt aaagcacgag gaagcggtca gcccattcgc
     5941 cgccaagctc ttcagcaata tcacgggtag ccaacgctat gtcctgatag cggtccgcca
     6001 cacccagccg gccacagtcg atgaatccag aaaagcggcc attttccacc atgatattcg
     6061 gcaagcaggc atcgccatgg gtcacgacga gatcctcgcc gtcgggcatg cgcgccttga
     6121 gcctggcgaa cagttcggct ggcgcgagcc cctgatgctc ttcgtccaga tcatcctgat
     6181 cgacaagacc ggcttccatc cgagtacgtg ctcgctcgat gcgatgtttc gcttggtggt
     6241 cgaatgggca ggtagccgga tcaagcgtat gcagccgccg cattgcatca gccatgatgg
     6301 atactttctc ggcaggagca aggtgagatg acaggagatc ctgccccggc acttcgccca
     6361 atagcagcca gtcccttccc gcttcagtga caacgtcgag cacagctgcg caaggaacgc
     6421 ccgtcgtggc cagccacgat agccgcgctg cctcgtcctg cagttcattc agggcaccgg
     6481 acaggtcggt cttgacaaaa agaaccgggc gcccctgcgc tgacagccgg aacacggcgg
     6541 catcagagca gccgattgtc tgttgtgccc agtcatagcc gaatagcctc tccacccaag
     6601 cggccggaga acctgcgtgc aatccatctt gttcaatcat atggtttcta tattatcttt
     6661 gtactaaaga gcaattgata atgtgcgaga aaaactggtc cttatatacc gtttgcagca
     6721 ctccctcccg aactttacga aaagtcgtgc gccacctgat tttcatcacg ccaaaaacct
     6781 acacgtatga ctactccggg ccagtgttca ccacgagcta tatagtgtta attaattacc
     6841 ttattggtta gctctgcatg taagggtggt gtgagccggg aattgggtct actctagcgt
     6901 tcagtaaggt gatataaagc tctgtagaat tcactggccg tcgttttaca acgtcgtgac
     6961 tgggaaaacc ctggcgttac ccaacttaat cgccttgcag cacatccccc tttcgccagc
     7021 tggcgtaata gcgaagaggc ccgcaccgat cgcccttccc aacagttgcg cagcctgaat
     7081 ggcgaatggc gcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc
     7141 atatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca gccccgacac
     7201 ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc cgcttacaga
     7261 caagctgtga ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa
     7321 cgcgcga
//
